Prostate cancer is one of the most common cancers in men and is one of the most common causes of cancer death, although, if diagnosed early, is potentially curable through surgical intervention, radiation therapy, or hormonal therapy. Since differential diagnosis is difficult at best, however, most prostate cancer is not diagnosed until later stages, typically after metastasis of the primary tumor. The primary therapy for metastatic prostate cancer is either androgen-antagonistic agents or castration, but most relapse patients show prostate tumor cells that are androgen-independent. Currently, there are no effective chemotherapeutic agents that can control the growth of androgen-independent prostate tumor cells.
PSA is a 240 amino acid member of the glandular kallikrein gene family. This 33 kDa single chain glycoprotein is a serine protease secreted by both normal and transformed epithelial cells of the prostate gland. PSA can be detected at low level in the sera of healthy males without evidence of prostate cancer. However, during neoplastic states, circulating levels of this antigen increase markedly, correlating with the clinical stage of the disease. PSA expression is almost exclusively restricted to the prostate cells, and is the most widely used marker for the diagnosis and monitoring of prostate cancer patients.
The tissue specificity of PSA makes it a particularly attractive target antigen for the development of immunotherapies against prostate cancer. PSA is detected in nearly all adenocarcinomas of the prostate and the expression of PSA was also demonstrated at distant metastatic sites. Several studies point out that PSA is a potential target for the induction of T-cell directed immunity against prostate cancer. Indeed, several PSA peptides capable of binding to MHC-class I molecules have been identified and it is now well established that a PSA specific T-cell repertoire exists in humans. Studies using in vitro immunization have shown the generation of CD4 and CD8 cells specific for PSA and T-cell lines with specific recognition of PSA peptides were generated from both healthy volunteer and prostate cancer patients. Of particular importance is the generation in vitro of cytotoxic T-lymphocytes specific for PSA and capable of lysing human prostate tumor cells [Xue et al., Prostate 30(2):73-8, (1997); Correale et al., Journal of the National Cancer Institute 89(4):293-300 (1997); Correale et al., Journal of Immunology 161(6):3186-94 (1998); Alexander et al., Urology 51(1):150-7 (1998)].
Based on these promising data, numbers of PSA-targeted immunotherapeutic approaches are currently being investigated in preclinical setting. Clinical trials employing a recombinant vaccinia virus engineered to express PSA, recombinant PSA encapsulated into liposomes, or autologous dendritic cells loaded with peptide sequences of PSA have already started [See Future Oncology Vol 4, No. 3/4 (1998)].
The present invention describes a new immunotherapeutic product for prostate cancer that employs as a therapeutic agent a binding agent such as a monoclonal antibody directed against PSA. It is believed that the therapeutic efficacy of the injected anti-PSA antibody is based on multiple mechanisms of actions acting in synergy. Tumor cell killing through an ADCC or CDC mechanism is not expected for these binding agents since PSA is not expressed at the cell surface. The therapeutic efficacy of these binding agents relies on the induction of a specific PSA cellular immune response and in the non-specific stimulation of the host immune system resulting in the induction of an immune response against various tumor antigens.
The induction of a specific cellular immune response upon immunization of the host with either Ab1 or Ab2 has been demonstrated in a number of studies. Of particular importance is the generation through this mechanism of specific CTLs responses in ovarian cancer patients, melanoma patients, myeloma patients, and non-Hodgkin's lymphoma patients [Nelson et al., Blood 88(2):580-9 (1996); Madiyalakan et al., Hybridoma 16(1):41-5 (1997); Osterborg et al., Blood 91 (7):2459-66 (1998); and Pride et al., Clinical Cancer Research 4:2363 (1998)]. It is therefore expected that the immunization of prostate cancer patients with the binding agents of the present invention may also induce a specific and protective CTL immune response against PSA.
This makes PSA an attractive target for immunotherapy. Several attempts at generating an immune response have met with limited success [Xue et al, The Prostate, 30:73-78 (1997); Correlae et al, J. National Cancer Institute, 89:293-300 (1997); Choe, et al, Cancer Investigations, 5:285-291 (1987); Wei, et al, Cancer Immunol. Immunother., 42:362-368 (1996), and International Application No. PCT/US97/04454, filed 19 Mar. 1997]. Combining the profound impact of prostate cancer with the lack of effective therapies, it is clear that alternative modalities of treatment need to be explored and that the ability to elicit a therapeutic immune response to PSA would be highly desirable.
Immunotherapies involve one or more components of the immune system to trigger a complex cascade of biological reactions focused on eliminating a foreign molecule from the host. The immune system consists of a wide range of distinct cell types, the most important of which are the lymphocytes. Lymphocytes determine the specificity of immunity, and it is their response that orchestrates the effector limbs of the immune system. Cells and proteins, such as antibodies, that interact with lymphocytes play critical roles in both the presentation of antigen and in the mediation of immunologic functions.
Individual lymphocytes provide a specialized function by responding to a limited set of structurally related antigens. As noted in more detail below, this function is defined structurally by the presence on the lymphocyte's surface membrane of receptors that are specific for binding sites (determinants or epitopes) on the antigen. Lymphocytes differ from each other not only in the specificity of their receptors, but also in their functions. One class of lymphocytes, B cells, are precursors of antibody-secreting cells, and function as mediators of the humoral immune response. Another class of lymphocytes, T cells, express important regulatory functions, and are mediators of the cellular immune response.
Cancer immunotherapy is based on the principle of inducing the immune system to recognize and eliminate neoplastic cells. The key elements in any immunotherapy is inducing the host immune system to first recognize a molecule as an unwanted target, and then inducing the system to initiate a response against that molecule. In healthy hosts, the immune system recognizes surface features of a molecule that is not a normal constituent of the host (i.e., is “foreign” to the host). Once the recognition function occurs, the host must then direct a response against that particular foreign molecule.
Both the recognition and the response elements of the immune system involve a highly complex cascade of biological reactions. In most immunologically based disorders, at least one of the steps in the recognition phase, or at least one of the steps in the response phase, are disrupted. Virtually any disruption in either of these complex pathways leads to reduced response or to no response. The inability of the immune system to destroy a growing tumor has been attributed, among other factors, to the presence of tumor-associated antigens (TAA) that induce immunological tolerance and/or immunosuppression. For example, in some kinds of cancer, the cancer itself tricks the host into recognizing a foreign cancer cell as a normal constituent, thus disrupting the recognition phase of the immune system. The immunological approach to cancer therapy involves modification of the host-tumor relationship so that the immune system is induced or amplifies its response to the TAAs. If successful, inducing or amplifying the immune system can lead to tumor regression, tumor rejection, and occasionally, to tumor cure.
One of the host system's mechanisms for combating a foreign molecule is called a humoral response, the production of an antibody against a specific foreign molecule (called an antigen). Typically, the antibody's capability of binding the antigen is based on highly complementary structures. That is, the shape of the antibody must contain structures that are the compliment of the structures on the antigen. When the respective structures are fully complimentary, then the two molecules bind tightly.
Antigens are molecules that interact with specific lymphocyte receptors—surface T cell antigen receptors and B cell immunoglobulin receptors. A particular B or T cell binds to a very specific region of the antigen, called an antigenic determinant or epitope. Thus antigens are molecules that bear one or more epitopes which may be recognized by specific receptors in an immune system, a property called antigenicity.
Immunogenicity is the property of stimulating the immune system to generate a specific response. Thus, all immunogens are antigens, but not vice-versa. Although an immune system may recognize an antigen, it does not respond to the antigen unless the antigen is also immunogenic.
An immune response to a particular antigen is greatly influenced by the structure and activity of the antigen itself, as well as myriad other factors. In some cases, the immune system is not able to generate an immune response to a particular antigen, a condition that is called tolerance.
In influencing whether an antigen is immunogenic or immunotolerant, an important characteristic of the antigen is the degree of difference between the antigen and similar molecules within the host. The most immunogenic antigens are those that have no homologs in the host, i.e., those that are most “foreign.” Other factors that promote immunogenicity include higher molecular weight, greater molecular complexity, the proper antigen dose range, the route of administration, the age of the host, and the genetic composition of the host.
As noted above, antigens may have one or more epitopes or binding sites that are recognized by specific receptors of the immune system. Epitopes may be formed by the primary structure of a molecule (called a sequential epitope), or may be formed by portions of the molecule separate from the primary structure that juxtapose in the secondary or tertiary structure of the molecule (called a conformational epitope). Some epitopes are hidden in the three dimensional structure of the native antigen, and become immunogenic only after a conformational change in the antigen provides access to the epitope by the specific receptors of the immune system. This is an important feature and function in the ability of a therapeutic reagent to initiate recognition and response to an antigen, the inducing both a cellular and humoral response to the antigen, and to increasing the antigenicity of a molecule without affecting its immunogenicity.
One of the responses generated by the immune system, a humoral response, involves the production of antibodies. Antibodies bear three major categories of antigen-specific determinants—isotypic, allotypic, and idiotypic—each of which is defined by its location on the antibody molecule. For the purpose of the present invention, we shall only focus on the idiotypic category.
Idiotypic determinants, or idiotopes, are markers for the V region of an antibody, a relatively large region that may include several idiotopes each capable of interacting with a different antibody. The set of idiotopes expressed on a single antibody V region constitutes the antibody idiotype. An antibody (Ab1) whose antigen combining site (paratope) interacts with an antigenic determinant on another antibody V region (idiotope) is called an anti-idiotypic antibody (Ab2). Thus, an antibody includes an antigen binding site, and may include one or more antibody binding sites. There are two types of anti-idiotypic antibodies, sometimes called Ab2α and Ab2β. In one type of anti-idiotype antibody (Ab2β), the combining site perfectly mimics the structure of the antigen epitope recognized by the Ab1 antibody. This type of anti-idiotype is said to represent the internal image of the antigen. By definition, the antigen and this type of anti-idiotype antibody compete for the same binding site on Ab1, and the antigen inhibits the interaction between Ab1 and the anti-idiotypic antibody. The phenomenon of producing an anti-idiotypic antibody having the internal image of the antigen may permit the use of antibodies to replace the antigen as an immunogen.
The second type of anti-idiotype, Ab2α, binds to an idiotope of Ab1 that is distinct from the antigen binding site, and therefore may be characterized in terms of the antigen's inability to prevent the binding of the anti-idiotype to Ab1. For this type of anti-idiotype, Ab1 can bind to both the antigen and the anti-idiotypic antibody. For a graphic representation of these types of antibodies and their interaction, see FIG. 1.
These various interactions based on idiotypic determinants is called the idiotypic network is based on the immunogenicity of the variable regions of immunoglobulin molecules (Ab1) which stimulate the immune system to generate anti-idiotypic antibodies (Ab2), some of which mimic antigenic epitopes (“internal image”) of the original antigen. The presence of internal image antibodies (Ab2) in the circulation can in turn induce the production of anti-anti-idiotypic antibodies (Ab3), some of which include structures that react with the original antigen.
In addition to a humoral response, the immune system may also generate a cellular response mediated by activated T-cells. There are a number of intercellular signals important to T cell activation. Under normal circumstances an antigen degrades or is cleaved to form antigen fragments or peptides. Presentation of antigen fragments to T-cells is the principal function of MHC molecules, and the cells that carry out this function are called antigen-presenting cells (APC: including but not limited to dendritic cells, macrophages, and B cells).
In addition to generating a humoral response, Ab1 and Ab2 have been shown to induce a cellular immune response characterized by proliferative lymphocytes (helper and suppressor lymphocytes), as well as cytotoxic lymphocytes. Therefore, according to the idiotypic network theory, the injection of anti-PSA antibody should result in the induction of a specific cellular and humoral immune response against the PSA molecule. The concept that anti-idiotypic antibodies function as immunogens has been shown by successful immunization against tumoral, bacterial, viral and parasitic antigens in animal models. Generating Ab2 is an indicator of the existence of a robust immune response that inherently reflects the induction of immune system pathways.
The capture and processing of an antigen by APCs is essential for the induction of a specific immune response. The three major APCs are dendritic cells, macrophages and B-lymphocytes; dendritic cells are the most efficient. The injected antibody forms a complex with a circulating PSA, and can be targeted to dendritic cells and macrophages through the Fc-receptors present on these cells. However the high number of Fc receptors on neutrophils may considerably limit this process.
The capture and processing of PSA by B-cells may also occur through the interaction of the membrane bound Ab2 with the anti-PSA/PSA complexes and in a similar manner through the interaction of membrane bound Ab3 with PSA (complexed or not with the anti-PSA antibody).